Multiplex PCR-based synchronous and rapid method for detecting 13 pathogenic microorganisms in water

A technology of pathogenic microorganisms and detection methods, which is applied in the determination/testing of microorganisms, biochemical equipment and methods, and resistance to vector-borne diseases, etc., and can solve the problem of lower sensitivity and specificity of primer target sequences, non-specific products, and amplification failure and other problems, to achieve the effect of simple operation, strong specificity and strong stability

Inactive Publication Date: 2012-10-03
LOGISTICAL ENGINEERING UNIVERSITY OF PLA +1
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Problems solved by technology

Although conventional PCR is a specific, sensitive, simple and rapid detection technology, it requires a long time for sample purification process, and once there is a very small amount of exogenous DNA contamination, false positive results may occur; multiple pairs of primers simultaneously Amplification, i

Method used

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  • Multiplex PCR-based synchronous and rapid method for detecting 13 pathogenic microorganisms in water
  • Multiplex PCR-based synchronous and rapid method for detecting 13 pathogenic microorganisms in water
  • Multiplex PCR-based synchronous and rapid method for detecting 13 pathogenic microorganisms in water

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Embodiment Construction

[0026] 1. Detection object

[0027] This method is mainly aimed at 13 kinds of pathogenic microorganisms that may exist in the water and may enter the water when public emergencies occur, namely: Escherichia coli, enterohaemorrhagic Escherichia coli O157:H7, Legionella pneumophila, Salmonella enteritidis , Shigella, Staphylococcus aureus, Listeria monocytogenes, Helicobacter pylori, Mycobacterium tuberculosis, Klebsiella pneumoniae, Vibrio cholerae, Bacillus anthracis, Yersinia pestis.

[0028] 2. Detection process

[0029] 2.1 Water sample treatment

[0030] Collect an appropriate amount of water samples, package them in sterile plastic bottles, and send them to the laboratory on the same day. After standing at room temperature for 1h~2h, take the supernatant and add it to the funnel of the filter funnel device of the Milliflex Plus microbial filtration system. Turn on the concentrator to concentrate and collect bacteria for 3 minutes. After the concentration is completed...

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Abstract

The invention relates to a multiplex PCR-based synchronous and rapid method for detecting 13 pathogenic microorganisms in water, which comprises the steps of using multiplex PCR to simultaneously amplify gene-specific fragments of the 13 pathogenic microorganisms including escherichia coli, enterohaemorragic escherichia coli o157:h7, legionella pneumophila, salmonella enteritidis, shigella dysenteriae, staphyloccocus aureus, listeria monoeytogenes, helicobacter pylori, mycobacterium tuberculosis, klebsiella pneumonia, vibrio cholera, bacillus anthracis and yersinia pestis, and detecting PCR amplified products through agarose gel electrophoresis, thereby achieving synchronous and rapid detection for the 13 pathogenic microorganisms.

Description

technical field [0001] The invention belongs to the technical fields of environmental science and engineering and environmental protection. Background technique [0002] At present, the methods commonly used to detect and identify pathogenic microorganisms at home and abroad mainly include traditional biochemical identification methods, immunological detection techniques, conventional PCR and other simple molecular biology and immunological methods. [0003] The traditional biochemical identification method (such as isolation culture, biochemical identification, etc.) is the most widely used, and it is the national standard testing procedure such as "Food Sanitation Microbiological Inspection (GB / T 4789-2003)". This method can obtain the number and characteristics of bacteria in the sample, etc. Qualitative and quantitative results are widely used due to simple operation, economy and good accuracy, but most of them need to go through 4 steps of pre-enrichment, enrichment, se...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12Q1/04C12Q1/14C12Q1/10
CPCY02A50/30
Inventor 方振东张春秀谢朝新敖漉王大勇马颖陈金丝
Owner LOGISTICAL ENGINEERING UNIVERSITY OF PLA
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